Equipment for fluorescence or luminescence detection, hereinafter also simply called fluorescence scanners, can be used to detect various molecular factors. Substances having different molecular properties can have different fluorescent properties, which can be detected in a targeted way. The fluorescence detection is optically based and hence is noninvasive or minimally invasive. With the knowledge of the applicable fluorescent properties, it is possible to ascertain the molecular nature of a given material being examined.
Besides the fluorescent properties, luminescent properties can also be analyzed in the same way. For the sake of simplicity, the term “fluorescence” will be used exclusively hereinafter, but it should be understood to mean both fluorescence and luminescence. Moreover, still other optical emissions phenomena based on comparable excitation processes are understood to be included in this term.
In both human and veterinary medicine, molecular properties, which may be termed a “molecular signature”, provide information about the state of health of a living creature or patient and can therefore be assessed diagnostically. Molecular signatures can be used in particular for detecting cancer. Still other syndromes, such as rheumatoid arthritis or arteriosclerosis of the carotid artery, can thus be identified.
Fluorescence may be excited by optical excitation. The excitation light can be in the infrared (IR) range, or in the near infrared range (NIR). The suitable optical frequency range is also dependent on the substance to be examined. Substances that themselves have no molecular or chemical properties that would be suitable for fluorescence detection can be molecularly “marked”. For example, markers that with suitable preparation bind to or to be deposited only on very special molecules can be used. Such marking functions by a mechanism that, in pictorial terms, can be thought of as a lock-and-key mechanism. The marker and the molecule to be detected fit one another like a lock and key, while the marker does not bind to other substances. If the marker has known fluorescent properties then, after the binding or deposition, the marker can be optically detected. The detection of the marker then allows conclusions to be drawn as to the presence of the marked special substance. For detection, accordingly, only one detector is needed, being capable of detecting light in the wavelength that the substance in question, or the marker used, emits upon excitation.
Such fluorescence methods may be used for examinations of regions near the surface or examinations in the open body (intra-operative applications). Examples of such investigations would be detecting fluorescently marked skin cancer or the detection of tumor boundaries in the resection of fluorescently marked tumors. For example, a system for making coronary arteries and the function of bypasses (that is, the blood flow through them) visible intra-operatively has been developed.
One subject of research in biotechnology is fluorescent metabolic markers that accumulate only in certain regions (such as tumors, infections, or other foci of disease), or are distributed throughout the body but are activated only in certain regions. Activation may be by, for example, tumor-specific enzyme activities or by additional exposure to light.
In medical diagnosis, marker substances, so-called fluorophores such as indocyanin green (ICG) are known which, for example, bind to blood vessels and can be detected optically. In an imaging process, the contrast with which blood vessels are displayed can be enhanced. So-called “smart contrast agents” are also becoming increasingly important. Activatable fluorescence markers may bind, for example, to tumor tissue, and the fluorescent properties are not activated until the binding to the substance to be marked occurs. Such substances may comprise self-quenched dyes, such as Cy5.5, which are bound to larger molecules by way of specific peptides. The peptides can in turn be detected by means of specific proteases, produced for example in tumors, and can be cleaved. The fluorophores are released by the cleavage and are no longer self-quenched but instead develop their fluorescent properties. The released fluorophores can be activated for example in the near IR wavelength range of around 740 nm. One example of a marker on this basis is AF 750 (Alexa Fluor 750), with a defined absorption and emission spectrum in the wavelength range of 750 nm (excitation) and 780 nm (emission).
In medical diagnosis, such activatable markers can be used for example for intra-operative detection of tumor tissue, so that the diseased tissue can be identified exactly and then removed. One typical application is the surgical treatment of ovarian cancer. Here the diseased tissue is typically removed surgically, and the patient later treated by chemotherapy. Because of the increased sensitivity of fluorescence detection, the diseased tissue can be better detected along with various surrounding foci of disease and thus removed more completely.
In the treatment of breast cancer, typical surgical treatments are lumpectomies (or mastectomies) and lymph node sections and lymph node biopsies. Lymph nodes are typically detected optically by means of 99mTc sulfur colloids in combination with low-molecular methylene blue. The radioactive mTc sulfur colloids could be avoided by using fluorescence detection, with correspondingly favorable effects on the health of the patient.
In the removal of brain tumors, the precise demarcation of the tumor tissue, which is attainable by the use of fluorescence detection, is of obvious importance. The treatment of pancreatic tumors can benefit from additional lymph node biopsies which could be identified by fluorescence detection, to detect possible intestinal cancer. In the treatment of skin cancer, the detection of skin neoplasms could be improved by fluorescence detection. The treatment of rheumatoid arthritic diseases of joints could improve medication monitoring in the sense that the extent of protease overproduction could be detected quantitatively, and the medication provided to counteract protease overproduction could be adapted quantitatively.
In treating these diseases which are identified as examples, as well as other syndromes, an operation may be performed in which the diseased tissue is removed surgically. Fluorescence detection can be performed to improve the detection of the diseased tissue portions to be removed during an ongoing operation in the open wound. To that end, the tissue parts must be marked before the operation with a suitable substance that is then activated by binding to the diseased tissue parts. An apparatus for fluorescence detection should therefore be easy for the surgeon to manipulate and should be usable in the sterile field of the operating room.
The detection of a region marked fluorescently in this way is done by exposing the region to light in the particular excitation wavelength of the fluorescent dye, and detecting the emitted light in the corresponding emission wavelength of the fluorophore. A fluorescence scan is made by recording a fluorescence image based on fluorescent light along with an optical image based on visible light. Next, the optical image and the fluorescence image are superimposed, in order to display the fluorescence in the context of the visual image. From the superimposed view (fusion) of the optical and fluorescence images on a display device, the surgeon can detect the tumor tissue and locate it in the body of the actual patient. The fused image with the fluorescently marked tissue is displayed on a screen on the fluorescence scanner or on an external computer with image processing software.
Typically, the excitation of the fluorescence of the marker is done by means of light, and the detection device must have a light source of adequate intensity, in order to penetrate the tissue to be examined to a depth of from 0.5 to 1 cm. In addition, an optical detector is necessary that on the one hand is capable of detecting the fluorescent light and on the other, if the fluorescent light is not in the visible wavelength range, also to record an image in the visible wavelength range.
For recording both an optical image and a fluorescence image, a beam splitter may be provided. The beam splitter splits the beam of light, arriving from the object or body to be examined, into one beam whose spectrum is in the IR or near IR (NIR) range of fluorescence and a further beam in the visible wavelength range. The IR or NIR beam is carried to an image sensor, provided specifically for it, and the visible beam is carried to an image sensor suitable for it. The two image sensors, separately from one another, simultaneously record an image. Thus, the fluorescence image and the optical image are available and can be superimposed on one another. It is a disadvantage that two image sensors are required, and that the construction is relatively bulky.
For recording both an optical image and a fluorescence image, a filter changer can be located in the beam path prior to the image sensor. The filter changer changes to a specific filter for taking fluorescence images and a specific filter for taking optical images. At least for recording fluorescence images, a change must be made to a filter that filters out light in the visible wavelength range, because otherwise the fluorescent light would be washed out by the visible light. A disadvantage is that the filter changer is mechanically complicated and makes for a bulky construction. In addition, the optical and the fluorescence image must be taken in succession, which makes the recording more time-consuming and tends to promote artifacts in the image caused by scanner motion between obtaining the images.
Existing apparatus for generating fluorescence scans is predominantly limited to the generation of 2D scans. It can be desirable to have 3D scans available as well. 3D information may be acquired by time-domain measuring methods, spectroscopically by laser projection, or by means of fluorescence mediated tomography. The known methods are complicated, however, because they are partly based on using still measuring methods that are highly computation-intensive.